1. Field of the Invention
The present invention relates to improved gene transfer methods and, more particularly, methods that enable highly efficient and widespread delivery of selected nucleic acids, to solid organs such as the heart or liver as well as to other solid cell masses such as a solid tumor. Preferred methods of the invention include treatment of tissue and/or cells for delivery of nucleic acid with one or more phosphodiesterase inhibitor compounds such as sildenafil.
2. Background
Effective delivery of nucleic acid to cells or tissue with high levels of expression are continued goals of gene transfer technology. As a consequence of the general inability to achieve those goals to date, however, clinical use of gene transfer methods has been limited.
Thus, for example, several delivery schemes have been explored for in vivo myocardial gene transfer, but none has proven capable of modifying a majority of cardiac myocytes in a homogeneous fashion. Techniques involving injection directly into the myocardium are considered of limited use because gene expression does not extend significantly beyond the needle track. R. J. Guzman et al. Circ Res 1993; 73:1202–1207; A. Kass-Eisler Proc Natl Acad Sci 1993; 90:11498–11502. In one study, percutaneous intracoronary delivery of 1010 pfu of adenovirus caused infection in only about one-third of the myocytes in the region served by the target artery. E. Barr et al. Gene Therapy 1994, 1:51–58.
Other coronary delivery models, either in situ or ex vivo, have produced a very small percentage of infected cells spread throughout the heart. J. Muhlhauser et al. Gene Therapy 1996; 3:145–153; J. Wang et al. Transplantation 1996; 61:1726–1729. To date, no in vivo delivery system has been able to infect a majority of cells in an intact heart.
Certain gene delivery procedures also have been quite invasive and hence undesirable. For example, one report describes essentially complete loss of endothelium by mechanical or proteolytic means to enable gene transfer from blood vessels to cells positioned across interposing endothelial layers. See WO 93/00051.
Certain gene transfer applications also have been explored in other organs such as the liver. In particular, ex vivo strategies have included surgical removal of selected liver cells, genetic transfer to the cells in culture and then reimplantion of the transformed cells. See M. Grossman et al., Nat Genet 1994, 6:335–341. Such an ex vivo approach, however, suffers from a number of drawbacks including, for example, the required hepatocyte transplantation. M. A. Kay et al., Science 1993, 262:117–119; and S. E. Raper et al., Cell Transplant 1993, 2:381–400. In vivo strategies for gene transfer to the liver also have been investigated, but have suffered from low delivery efficiencies as well as low specificity to the targeted tissue. N. Ferry et al., Proc Natl Acad Sci USA 1991, 88:8377–8391; A. Lieber et al. Proc Natl Acad Sci USA 1995, 6:6230–6214; A. L. Vahrmeijer et al., Reg Cancer Treat 1995, 8:25–31. See also P. Heikkilia et al., Gene Ther 1996, 3(1):21–27.
Gene transfer has been generally unsuccessful in additional applications. For example, gene transfer therapies for treatment of cystic fibrosis have largely failed because transduction of insufficient numbers of cells.
It thus would be desirable to have improved methods and systems to effectively deliver nucleic acid to targeted cells and tissue. It would be particularly desirable to have new methods and systems for effective delivery of nucleic acids into solid organs, especially the heart, liver, lung and the like, as well as other solid cell masses such as a solid tumor.